CN102621265A - Method for measuring contents of multiple components in Shenxiong glucose injection - Google Patents

Method for measuring contents of multiple components in Shenxiong glucose injection Download PDF

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CN102621265A
CN102621265A CN2012100844792A CN201210084479A CN102621265A CN 102621265 A CN102621265 A CN 102621265A CN 2012100844792 A CN2012100844792 A CN 2012100844792A CN 201210084479 A CN201210084479 A CN 201210084479A CN 102621265 A CN102621265 A CN 102621265A
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glucose injection
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chuanxiong
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CN102621265B (en
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叶湘武
郑林
王永林
兰燕宇
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GUIZHOU JINGFENG INJECTION CO Ltd
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Abstract

The invention belongs to the technical field of quality control of Chinese materia medica preparations, and provides a method for measuring contents of multiple components in Shenxiong glucose injection. According to the invention, the HPLC (high performance liquid chromatography) is adopted to measure contents of six compounds, namely tanshinol, ligustrazine hydrochloride, protocatechuic acid, rosmarinic acid, danshinolic acid B and danshinolic acid A, in the Shenxiong glucose injection under the same chromatographic condition. Compared with the present simple content measurement method for a single component, the method provided by the invention is better in integrity, characteristics and stability, can comprehensively reflect the interior quality of the preparations, and provides test base for further improving the quality control level of the Shenxiong glucose injection and ensuring the quality stability of the products.

Description

The multicomponent content assaying method of ginseng rhizome of chuanxiong glucose injection
Technical field
The present invention relates to the multicomponent content assaying method of a seed ginseng rhizome of chuanxiong glucose injection, belong to the technical field of Chinese medicine preparation quality control.
Background technology
Ginseng rhizome of chuanxiong glucose injection is made up of the red sage root and Ligustrazine Hydrochloride compatibility, has platelet aggregation-against, coronary artery dilator effect, the clinical treatment that is used for obliterated cerebral vascular disease and other ischemic angiopathy.The assay project of the ginseng existing quality standard of rhizome of chuanxiong glucose injection (WS-10001-(HD-1136)-2002) is only quantitatively controlled danshensu and Ligustrazine Hydrochloride, is difficult to reflect the chemical constitution characteristic of preparation comprehensively.Along with the raising of China's pharmaceutical production quality control level and deepening continuously to injection safety, curative effect, problem attention degree such as quality controllable; State Food and Drug Administration has proposed " national drug standards raising action plan " and " the traditional Chinese medicine security revalues work " in succession, with comprehensive lifting drug quality controlling level.Therefore, set up a kind of ability and control ginseng rhizome of chuanxiong glucose injection method for quality fast, accurate and comprehensively, realize comprehensive control, have great importance its material crowd.
Summary of the invention
The objective of the invention is to, the multicomponent content assaying method of a seed ginseng rhizome of chuanxiong glucose injection is provided.
The present invention is directed to the simple content assaying method of existing single component; Propose first ginseng rhizome of chuanxiong glucose injection is carried out multicomponent assay simultaneously; To join in the rhizome of chuanxiong glucose injection that content is higher, chromatographic resolution preferably danshensu, Ligustrazine Hydrochloride, protocatechualdehyde, Rosmarinic acid, tanshin polyphenolic acid B, 6 kinds of chemical constitutions of salviandic acid A as the assay parameter; Make the content assaying method of ginseng rhizome of chuanxiong glucose injection have more globality, characteristic and stability; And instrument, chromatographic column, detecting device, detection wavelength, chromatogram flow phase, column temperature etc. are optimized; Through the quantitative control of multi-target ingredient, can concentrated expression the inherent quality of ginseng rhizome of chuanxiong glucose injection, promote the quality control level of preparation.
Multicomponent content assaying method of the present invention is under same chromatographic condition, simultaneously to joining the high performance liquid chromatography assay of danshensu contained in the rhizome of chuanxiong glucose injection, Ligustrazine Hydrochloride, protocatechualdehyde, Rosmarinic acid, tanshin polyphenolic acid B, 6 kinds of chemical constitutions of salviandic acid A.
Multicomponent content assaying method of the present invention is made up of following steps:
One, chromatographic condition and system suitability test
Instrument: supper-fast liquid chromatographic system (UFLC, Tianjin, island comprise binary gradient pump, vacuum degassing machine, automatic sampler, column oven, PDAD, Lcsolution workstation);
Chromatographic column: Diamonsil C 18Chromatographic column (4.6mm * 250mm, 5 μ m);
Moving phase: A is an acetonitrile, and B is 0.05%H 3PO 4, carry out gradient elution by the regulation in the table 1;
Flow velocity: 1mL/min;
Column temperature: 40 ℃;
Detect wavelength: 220nm;
Sample size: 10 μ L.
Table 1 eluent gradient wash-out table
Figure BDA0000147496500000021
Two, the preparation of reference substance solution respectively precision to take by weighing Sodium Danshensu, Ligustrazine Hydrochloride, protocatechualdehyde, Rosmarinic acid, tanshin polyphenolic acid B, salviandic acid A an amount of, place 6 10mL volumetric flasks respectively, add dissolve with methanol and be diluted to scale; Shake up, promptly get mass concentration and be respectively 1.056,7.649; 1.084; 1.020,1.046,1.036mgmL -1The reference substance storing solution.Precision is measured above-mentioned 6 kinds of reference substance storing solutions 8,6,2,1.2,1.2 respectively, and 1.2mL puts in the same 20mL volumetric flask, adds methanol constant volume to scale, shakes up, and must mix the reference substance intermediate liquid.Precision is measured and is mixed reference substance intermediate liquid 0.25,0.5,1,2,4, and 8mL places 6 10mL volumetric flasks respectively, adds methanol constant volume to scale, shakes up, and promptly gets.
Three, the preparation precision of need testing solution is measured these article 10mL, adds the dilution of 5% glucose solution, is settled to 25mL, shakes up and promptly gets need testing solution.
Four, determination method is measured according to the chromatographic condition under the multicomponent content assaying method according to the invention, promptly gets.
This research is based on ginseng rhizome of chuanxiong glucose injection UPLC-Q-TOF mass spectrometry analysis and form the chemical constitution of the flavour of a drug red sage root of this parenteral solution compound; Finally confirmed in the ginseng rhizome of chuanxiong glucose injection index components of 6 kinds of main chemical compositions, set up corresponding multicomponent content assaying method as quality control.
Multicomponent assay research method of the present invention is the preferred plan that obtains through a large amount of shaker tests, and following experimental study is a preferred process of the present invention:
One, chromatographic condition and system suitability test
1, instrument and reagent
Supper-fast liquid chromatographic system (UFLC, Tianjin, island comprise binary gradient pump, vacuum degassing machine, automatic sampler, column oven, PDAD, Lcsolution workstation); AE2401/10 ten thousand electronic balances (plum Teller-Tuo benefit instrument Shanghai company limited); Ultrapure water machine (Sichuan Water Technology Development Co., Ltd.).Acetonitrile (German MERCK company) is a chromatographically pure; Water is ultrapure water; It is pure that other reagent are analysis.Sodium Danshensu (lot number 110855200508), Ligustrazine Hydrochloride (lot number 110817-200305), protocatechualdehyde (lot number 110810-200705) reference substance is purchased in Nat'l Pharmaceutical & Biological Products Control Institute; Rosmarinic acid (lot number MUST-10120901), tanshin polyphenolic acid B (lot number MUST-11031401), salviandic acid A (lot number MUST-10012901) reference substance are purchased in Beijing permanent unit and are opened a day chemical industry Institute for Research and Technology; Ginseng rhizome of chuanxiong glucose injection (Guizhou Jingfeng Injection Co., Ltd. provides).
2, the selection of chromatographic condition
Test was once investigated different proportioning flow phase system such as methanol-water, acetonitrile-water, acetonitrile-0.05% phosphoric acid, acetonitrile-0.5% acetate; Compare through degree such as grade or gradient test; The result shows; With acetonitrile-0.05% phosphoric acid is that eluent gradient wash-out separating effect is better, and 6 composition chromatographic peaks that detected can reach baseline separation, have shortened analysis time again.Respectively 6 kinds of compositions are carried out full wavelength scanner with PDAD and learn that the maximum absorption wavelength of danshensu, Ligustrazine Hydrochloride, protocatechualdehyde, Rosmarinic acid, tanshin polyphenolic acid B, salviandic acid A is respectively 221,215; 230; 219,221,221nm; Be detection signal strength and the detection sensitivity of taking into account each component, select 220nm as detecting wavelength.Through instrument, moving phase, detection wavelength, column temperature isochromatic spectrum condition are optimized on the basis of investigation, obtained to be applicable to the chromatographic condition of multicomponent assay:
Instrument: supper-fast liquid chromatographic system (UFLC, Tianjin, island comprise binary gradient pump, vacuum degassing machine, automatic sampler, column oven, PDAD, Lcsolution workstation);
Chromatographic column: Diamonsil C 18Chromatographic column (4.6mm * 250mm, 5 μ m);
Moving phase: A is an acetonitrile, and B is 0.05%H 3PO 4, carry out gradient elution by the regulation in the table 2;
Flow velocity: 1mL/min;
Column temperature: 40 ℃;
Detect wavelength: 220nm;
Sample size: 10 μ L.
Table 2 eluent gradient wash-out table
Figure BDA0000147496500000041
Two, the preparation of reference substance solution:
It is an amount of that precision takes by weighing Sodium Danshensu, Ligustrazine Hydrochloride, protocatechualdehyde, Rosmarinic acid, tanshin polyphenolic acid B, salviandic acid A respectively, places 6 10mL volumetric flasks respectively, adds dissolve with methanol and be diluted to scale; Shake up, promptly get mass concentration and be respectively 1.056,7.649; 1.084; 1.020,1.046,1.036mgmL -1The reference substance storing solution.Precision is measured above-mentioned 6 kinds of reference substance storing solutions 8,6,2,1.2,1.2 respectively, and 1.2mL puts in the same 20mL volumetric flask, adds methanol constant volume to scale, shakes up, and must mix the reference substance intermediate liquid.Precision is measured and is mixed reference substance intermediate liquid 0.25,0.5,1,2,4, and 8mL places 6 10mL volumetric flasks respectively, adds methanol constant volume to scale, shakes up, and promptly gets.
Three, the preparation of need testing solution:
Precision is measured these article 10mL, adds the dilution of 5% glucose solution, is settled to 25mL, shakes up and promptly gets need testing solution.
Four, the drafting of typical curve
Precision is drawn reference substance series solution respectively, injects UFLC by the chromatographic condition of drafting and measures, and with peak area the sample introduction quality is carried out linear regression, gets regression equation, related coefficient and the range of linearity of each tested composition, sees table 3.
The linear relationship of table 36 a tested composition
Figure BDA0000147496500000051
Five, precision test
By chromatographic condition under the multicomponent content assaying method according to the invention, precision is measured ginseng rhizome of chuanxiong glucose injection need testing solution 10 μ L continuous sample introductions 5 times, measures 6 kinds of compounds respectively, makes the peak area integration and calculates RSD, sees table 4.Wherein, A is a Sodium Danshensu, and B is a Ligustrazine Hydrochloride, and C is a protocatechualdehyde, and D is a Rosmarinic acid, and E is a tanshin polyphenolic acid B, and F is a salviandic acid A.Can know that by the result precision of multicomponent content assaying method according to the invention is good.
Table 4 Precision test result
Figure BDA0000147496500000061
Six, replica test
Get same lot number ginseng rhizome of chuanxiong glucose injection, the preparation method of shining need testing solution under the multicomponent content assaying method according to the invention prepares 6 parts of need testing solutions, measures respectively, calculates content and RSD, sees table 5.Wherein, A is a Sodium Danshensu, and B is a Ligustrazine Hydrochloride, and C is a protocatechualdehyde, and D is a Rosmarinic acid, and E is a tanshin polyphenolic acid B, and F is a salviandic acid A.Can know that by the result repeatability of multicomponent content assaying method according to the invention is good.
Table 5 replica test result
Figure BDA0000147496500000062
Seven, stability test
Get same ginseng rhizome of chuanxiong glucose injection need testing solution, respectively at 0,2,4,6, the 12h sample introduction is measured, and makes the peak area integration and calculates RSD, sees table 6.Wherein, A is a Sodium Danshensu, and B is a Ligustrazine Hydrochloride, and C is a protocatechualdehyde, and D is a Rosmarinic acid, and E is a tanshin polyphenolic acid B, and F is a salviandic acid A.Can know that by the result need testing solution of multicomponent content assaying method according to the invention is stable in 12h.
Table 6 stability test result
Figure BDA0000147496500000071
Eight, recovery test
Precision is measured totally 6 parts of the ginseng rhizome of chuanxiong glucose injection 5mL of same lot number; Put respectively in the 25mL measuring bottle; It is an amount of that accurate adding mixes reference substance solution, by preparation method's preparation of need testing solution under the multicomponent content assaying method according to the invention, measures by the chromatographic condition after optimizing under the multicomponent content assaying method according to the invention; Calculate recovery rate and RSD see table 7.Wherein, A is a Sodium Danshensu, and B is a Ligustrazine Hydrochloride, and C is a protocatechualdehyde, and D is a Rosmarinic acid, and E is a tanshin polyphenolic acid B, and F is a salviandic acid A.Can know that by the result recovery of multicomponent content assaying method according to the invention is good.
Table 7 recovery test result
Figure BDA0000147496500000081
Nine, sample size is measured
Get the ginseng rhizome of chuanxiong glucose injection of different batches; Each sample is parallel does 2 parts; Preparation method according to need testing solution under the multicomponent content assaying method according to the invention prepares need testing solution; By the content of 6 kinds of chemical constitutions of the mensuration of the chromatographic condition under the multicomponent content assaying method according to the invention, see table 8.Wherein, A is a Sodium Danshensu, and B is a Ligustrazine Hydrochloride, and C is a protocatechualdehyde, and D is a Rosmarinic acid, and E is a tanshin polyphenolic acid B, and F is a salviandic acid A.
Mensuration result (n=2) mgL of 6 kinds of chemical constitutions in the table 8 ginseng rhizome of chuanxiong glucose injection -1
Figure BDA0000147496500000091
Can know that by the result content basically identical of 6 contained tested compositions explains that the quality conformance of product is good between different batches in each batch ginseng rhizome of chuanxiong glucose injection.In 5 batches of samples: the average content of Sodium Danshensu is 209.004mgL -1, the average content of Ligustrazine Hydrochloride is 1071.139mgL -1, the average content of protocatechualdehyde is 21.454mgL -1, the average content of Rosmarinic acid is 29.630mgL -1, the average content of tanshin polyphenolic acid B is 35.299mgL -1, the average content of salviandic acid A is 21.020mgL -1
Through comprehensive evaluation; In conjunction with each system suitability parameter situation; And through the methodology checking, indexs such as this method degree of accuracy, repeatability, stability, the recovery are all good, through the quantitative control of multi-target ingredient; Inherent quality that can the concentrated expression preparation can be used for joining the quality control of rhizome of chuanxiong glucose injection.
Description of drawings
Fig. 1: Sodium Danshensu reference substance solution ultraviolet spectrogram.
Fig. 2: Ligustrazine Hydrochloride reference substance solution ultraviolet spectrogram.
Fig. 3: protocatechualdehyde reference substance solution ultraviolet spectrogram.
Fig. 4: Rosmarinic acid reference substance solution ultraviolet spectrogram.
Fig. 5: tanshin polyphenolic acid B reference substance solution ultraviolet spectrogram.
Fig. 6: salviandic acid A reference substance solution ultraviolet spectrogram.
Fig. 7: Sodium Danshensu, Ligustrazine Hydrochloride, protocatechualdehyde, Rosmarinic acid, tanshin polyphenolic acid B, salviandic acid A reference substance solution UFLC chromatogram (220nm).Chromatographic peak is corresponding respectively: A is a Sodium Danshensu, and B is a Ligustrazine Hydrochloride, and C is a protocatechualdehyde, and D is a Rosmarinic acid, and E is a tanshin polyphenolic acid B, and F is a salviandic acid A.
Fig. 8: need testing solution UFLC chromatogram (220nm).Chromatographic peak is corresponding respectively: A is a Sodium Danshensu, and B is a Ligustrazine Hydrochloride, and C is a protocatechualdehyde, and D is a Rosmarinic acid, and E is a tanshin polyphenolic acid B, and F is a salviandic acid A.
Embodiment:
Enumerate embodiment further explain the present invention below, this embodiment only is used to the present invention is described and the present invention is not limited.
Embodiment 1: the high performance liquid chromatography assay of joining Sodium Danshensu contained in the rhizome of chuanxiong glucose injection, Ligustrazine Hydrochloride, protocatechualdehyde, Rosmarinic acid, tanshin polyphenolic acid B, 6 kinds of chemical constitutions of salviandic acid A
One, chromatographic condition and system suitability test
Instrument: supper-fast liquid chromatographic system (UFLC, Tianjin, island comprise binary gradient pump, vacuum degassing machine, automatic sampler, column oven, PDAD, Lcsolution workstation);
Chromatographic column: Diamonsil C 18Chromatographic column (4.6mm * 250mm, 5 μ m);
Moving phase: A is an acetonitrile, and B is 0.05%H 3PO 4, carry out gradient elution by the regulation in the table 1;
Flow velocity: 1mL/min;
Column temperature: 40 ℃;
Detect wavelength: 220nm;
Sample size: 10 μ L.
Table 1 eluent gradient wash-out table
Figure BDA0000147496500000101
Figure BDA0000147496500000111
Two, the preparation of reference substance solution respectively precision to take by weighing Sodium Danshensu, Ligustrazine Hydrochloride, protocatechualdehyde, Rosmarinic acid, tanshin polyphenolic acid B, salviandic acid A an amount of, place 6 10mL volumetric flasks respectively, add dissolve with methanol and be diluted to scale; Shake up, promptly get mass concentration and be respectively 1.056,7.649; 1.084; 1.020,1.046,1.036mgmL -1The reference substance storing solution.Precision is measured above-mentioned 6 kinds of reference substance storing solutions 8,6,2,1.2,1.2 respectively, and 1.2mL puts in the same 20mL volumetric flask, adds methanol constant volume to scale, shakes up, and must mix the reference substance intermediate liquid.Precision is measured and is mixed reference substance intermediate liquid 0.25,0.5,1,2,4, and 8mL places 6 10mL volumetric flasks respectively, adds methanol constant volume to scale, shakes up, and promptly gets.
Three, the preparation precision of need testing solution is measured these article 10mL, adds the dilution of 5% glucose solution, is settled to 25mL, shakes up and promptly gets need testing solution.
Four, determination method is measured according to chromatographic condition under the multicomponent content assaying method according to the invention, promptly gets.

Claims (5)

1. join the multicomponent content assaying method of rhizome of chuanxiong glucose injection; It is characterized in that: said multicomponent content assaying method is under same chromatographic condition, simultaneously contained 6 kinds of chemical constitutions of danshensu, Ligustrazine Hydrochloride, protocatechualdehyde, Rosmarinic acid, tanshin polyphenolic acid B, salviandic acid A in the ginseng rhizome of chuanxiong glucose injection is carried out the liquid chromatography assay.
2. according to the multicomponent content assaying method of the described ginseng rhizome of chuanxiong of claim 1 glucose injection, it is characterized in that said multicomponent content assaying method is undertaken by following step:
(1) chromatographic condition: chromatographic column is the C18 post; Acetonitrile-0.05%H3PO4 solution gradient wash-out; Column temperature is 35 ℃~55 ℃; The detection wavelength is 200~400nm;
(2) preparation of reference substance solution: take by weighing Sodium Danshensu, Ligustrazine Hydrochloride, protocatechualdehyde, Rosmarinic acid, tanshin polyphenolic acid B, salviandic acid A reference substance respectively, add dissolve with methanol, get the reference substance storing solution; Precision is measured above-mentioned reference substance storing solution and is added methyl alcohol dilution respectively, reference substance solution;
(3) preparation of need testing solution: measure ginseng rhizome of chuanxiong glucose injection, add 5% glucose solution, promptly get need testing solution;
(4) determination method: measure according to the said chromatographic condition of step (1), promptly get.
3. according to the multicomponent content assaying method of the described ginseng rhizome of chuanxiong of claim 2 glucose injection, it is characterized in that: described step (1) chromatographic condition and system suitability test:
Instrument: supper-fast liquid chromatographic system (UFLC, Tianjin, island comprise binary gradient pump, vacuum degassing machine, automatic sampler, column oven, PDAD, Lcsolution workstation);
Chromatographic column: Diamonsil C18 chromatographic column (4.6mm * 250mm, 5 μ m);
Moving phase: A is an acetonitrile, and B is 0.05%H3PO4, carries out gradient elution by the regulation in the table 1;
Flow velocity: 1mL/min;
Column temperature: 40 ℃;
Detect wavelength: 220nm;
Sample size: 10 μ L.
Table 1 eluent gradient wash-out table
Figure FDA0000147496490000021
4. according to the multicomponent content assaying method of the described ginseng rhizome of chuanxiong of claim 2 glucose injection, it is characterized in that: the preparation of described step (2) reference substance solution:
It is an amount of that precision takes by weighing Sodium Danshensu, Ligustrazine Hydrochloride, protocatechualdehyde, Rosmarinic acid, tanshin polyphenolic acid B, salviandic acid A respectively, places 6 10mL volumetric flasks respectively, adds dissolve with methanol and be diluted to scale; Shake up, promptly get mass concentration and be respectively 1.056,7.649; 1.084; 1.020,1.046, the reference substance storing solution of 1.036mgmL-1.Precision is measured above-mentioned 6 kinds of reference substance storing solutions 8,6,2,1.2,1.2 respectively, and 1.2mL puts in the same 20mL volumetric flask, adds methanol constant volume to scale, shakes up, and must mix the reference substance intermediate liquid; Precision is measured and is mixed reference substance intermediate liquid 0.25,0.5,1,2,4, and 8mL places 6 10mL volumetric flasks respectively, adds methanol constant volume to scale, shakes up, and promptly gets.
5. according to the multicomponent content assaying method of the described ginseng rhizome of chuanxiong of claim 2 glucose injection; It is characterized in that: the preparation of described step (3) need testing solution: precision is measured ginseng rhizome of chuanxiong glucose injection 10mL; Add the dilution of 5% glucose solution; Be settled to 25mL, shake up and promptly get need testing solution.
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CN103149310B (en) * 2013-01-21 2015-05-20 贵州景峰注射剂有限公司 Fingerprint building method of Shenxiong glucose injection preparation
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CN105510472A (en) * 2015-12-22 2016-04-20 贵州景峰注射剂有限公司 Method for detecting tannin in shenxiong glucose injection
CN105628696A (en) * 2015-12-28 2016-06-01 上海景峰制药有限公司 Method for determining content of total phenolic acid in salivae miltiorrhizae liguspyragine hydrochloride and glucose injection
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